To study the cascade mechanism control of glutamine synthetase (GS) under conditions that will approximate the in vivo situation, Escherichia coli cells have been made permeable to only small molecules. The GS activity in permeabilized cells (but not in untreated cells) can be measured in situ by means of the same procedure used for measuring activity of soluble enzyme. In addition to GS, the permeabilized cells retain all the protein components involved in modification of GS and therefore are able to catalyze the adenylylation and deadenylyation of GS in situ. This interconversion responds to alterations in concentration of ATP, UTP, Pi Mg ions and allosteric effectors (alpha-ketoglutarate and glutamine) as predicted by the earlier in vitro studies with cell-free systems.